A series of N2-substituted guanine derivatives was screened against mammalian thymidine kinase and the thymidine kinase encoded by type I herpes simplex virus to examine their capacity to selectivity inhibit the viral enzyme. Several bases, nucleosides, and nucleotides displayed selective activity. The mechanism of action of the most potent derivative, N2-phenyl-2'-deoxyguanosine (PhdG) was studied in detail. PhdG (a) inhibited the viral enzyme competitively with respect to the substrates thymidine and deoxycytidine, (b) was completely resistant to phosphorylation, (c) displayed limited toxicity for the HeLa cell lines employed as hosts for viral infection, and (d) selectively inhibited viral thymidine kinase function in intact cultured cells. The results indicate that the PhdG drug prototype has potential as a selective anti-herpes agent and as a novel molecular probe of the structure and function of herpes simplex thymidine kinase.
The anomeric specificity of Escherichia coli CMP-N-acetylneuraminic acid (CMP-NeuAc) synthetase was investigated by NMR using 13C-labeled N-acetylneuraminic acid (NeuAc). Consumption of the beta-anomer of [2-13C]N-acetylneuraminic acid was observed upon addition of enzyme, with a concomitant appearance of an anomeric resonance for CMP-N-acetylneuraminic acid. Inhibition by substrate analogues the anomeric oxygen was determined in a similar manner using [2-13C,(50 atom %)18O]N-acetylneuraminic acid. An upfield shift of 1.5 Hz in the anomeric resonance of both the [13C]NeuAc substrate and CMP-[13C]NeuAc product was observed due to the 18O substitution. This result implies conservation of the NeuAc oxygen. Results of steady-state kinetic analysis suggest a sequential-type mechanism and therefore no covalent intermediate. Thus, CMP-beta-NeuAc is probably formed by a direct transfer of the anomeric oxygen of beta-NeuAc to the alpha-phosphate of CTP.
on a column of silica gel (eluting solvent: benzene). The first materia] to emerge from the column was the desired 2,8,17-trithia [45,12] [9]metacyclophane, which was recrystallized from ethyl acetate (85 mg, 0.27 mmol, 5.5% yield). The melting behavior was unusual: at 170-180 °C, the crystals softened and bent, but did not liquefy; at 280 °C the material began to discolor; decomposition was gradual from 300-350 °C (lit.1 2mp 176 °C). lH
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